Summary Staphylococcus aureus in an important Gram positive human pathogen that is capable of infecting numerous bodily sites including the lung. It leads to significant morbidity and mortality, has a high incidence of antibiotic resistance and is a pathogen identified as needing increased research to discovery new therapeutics. The long term goal of our research is to better understand the host-pathogen interaction between S. aureus and the host immune system. It is hoped that an improved understanding of this interaction could lead to novel therapies targeting the bacterium or modulating the host to thwart this multidrug resistant pathogen. The objective of this proposal is to characterize the innate immune response of the human lung as well as the initial response of S. aureus to the airway and identify the genetic factors important for causing infection. The rationale for this approach is that new targets are needed to either develop antibiotics/inhibitors or vaccines against S. aureus. This is due to its high level of antibiotic resistance and previously promising vaccine candidates in murine models have failed to translate to efficacious human vaccines. Our preliminary data shows that humanized mice represent an improved model for the study of S. aureus infection in the lung, allowing the role of human specific virulence factors to be studied and understand the immune response to infection. We present a novel model of humanized mice whereby mice are implanted with human lung tissue. We have also demonstrated the use of a Tn-seq library to identify new factors important for infection in the airway. This contribution is significant as it will provide an understanding of how the human immune system, including the epithelium responds to S. aureus, as well as how S. aureus reacts to this environment and the factors that it employs to sustain infection. The innovation of this research is the utilization of a new model of infection that leverages our experience with humanized models of infection. This model incorporates human lung tissue to better replicate the human lung. We will characterize this response at multiple levels, at the transcriptional, protein and cellular level. Global approaches to understand the pathogen response, RNA-seq and Tn-seq will be combined to identify new genetic elements important in pathogenesis. Given present failure with vaccines and current drug pipelines, investigating gene products in the context of human model outside of ell-characterized virulence factors might lead to novel discoveries.